The inhibition is Ag specific and requires the presence of the first cohort of T cells. (A) The inhibition depends on the presence of the first cohort of T cells. Naive (10⁶) or memory (2 × 10⁶) Marilyn T cells were injected into OT-II hosts that had been primed 6 days earlier with H-Y peptide–loaded mature DCs, in the presence (iii-vi) or not (i,ii) of a first cohort of Marilyn-Lat-DTR T cells. The day before the injection of the second cohort, Marilyn-Lat-DTR cells were deleted (v,vi) or not (iii,iv) by the injection of DT. Dot plots of gated CD4⁺CD45.1⁺ cells from the DLN studied 6 days later (day 6 + 6) are representative of at least 2 experiments with 2 mice each per group. (B) The inhibition is Ag specific. CD45.1 B6 mice were injected or not (i-iv) with a first cohort of naive CFSE-labeled Marilyn (v-viii) or OT-II (ix-xii) Tg CD4 T cells and were then immunized with LPS-matured DCs pulsed with both OVA and H-Y peptides. Six days later, CFSE-labeled naive (top panels) or memory (bottom panels) OT-II and Marilyn cells were coinjected in the same mouse. Proliferation was measured on the second cohorts (CD45.2⁺, and Vβ6⁺ for Marilyn, Vα2^hi for OT-II) in the same mouse 6 days later. Representative of 2 independent experiments with 2 mice each per group.